Breast cancer is the most commonly diagnosed form of cancer in women 40-55 years of age and it is the second major cause of cancer deaths behind lung cancer for all women. Metastatic breast cancer, where cancer cells spread by motile mechanisms and establish tumors at distant vital sites, is much harder to eradicate and is the primary cause of patient death from breast cancer. Understanding the molecular principles that determine the efficiency of tumor metastasis is therefore critical to the prevention and treatment of breast tumors. Traditional cancer therapeutics are aimed at preventing tumorigenesis of normal breast tissue and inhibiting growth of established cancers. However, few therapeutic strategies target cell migration and invasion, although the pathological deregulation of these processes is a major cause of morbidity associated with the disease. Cell migration and invasion are coordinately regulated by the small GTPase Rac1 and the localized production of the lipid phosphatidylinositol-4,5-bisphosphate (PI4,5P2). The hyperactivation of Rac1 signaling has been observed in many cancers, particularly in cancers of the breast, and this is directly linked to increased metastatic potential and poor patient survival. A role for PI4,5P2 signaling in cancer progression has so far not been reported. However, recent evidence described in the preliminary studies section of this proposal has established that PIPK1a, a member of the Type I phosphatidylinositol-4-phosphate kinase family, which generates PI4,5P2, is a critical regulator of cell migration and cell-matrix adhesion. We have defined a biochemical pathway in which PIPK1a mediates Rac1 activation in response to integrin and growth factor signals. Rac1, in turn, controls signaling to downstream effectors, including a second member of the PIPKI family, PIPK1b, to promote the assembly of F-actin and of focal adhesion sites necessary for migration and invasion. These results therefore establish a pathway in which PIPK1a is the pinnacle of a signaling cascade that links transmembrane receptors to the regulation of actin and focal adhesion assembly during cell motility. Because cell migration and adhesion are critical for cancer metastasis, PIPK1a may be a target for the prevention of cancer progression. The long-term goal of these studies is to validate PIPK1a as a target for therapeutic intervention in metastatic disease using tissue culture cell models and the athymic nude mouse model of breast cancer. The proposed research also involves pilot studies designed to assess the efficacy of a newly identified natural small-molecule inhibitor of PIPK1a in the control of breast cancer progression. We will use a combination of basic research, chemical genetic and in vivo approaches to systematically address the role of the PIPK1a pathway in cell migration and invasion in a 3-dimensional matrix, in anchorage-independent growth, and in cancer progression in vivo using the athymic nude mouse. The proposed research not only has the potential to impact therapeutic design to prevent breast cancer metastasis, but will also advance our understanding of signaling mechanisms that may be critical for breast cancer metastasis. PUBLIC HEALTH RELEVANCE: This research project employs cancer cell lines and mouse models of cancer metastasis to uncover the signaling mechanisms that control a cell's ability to move. The migration of cells is important for proper tissue formation, immune function and wound repair, but when aberrantly regulated can also form the basis of devastating human diseases including cancer, atherosclerosis and allergies. Our long-term goal is to better understand the signaling mechanisms that control cell migration and to use this information to develop new therapeutic approaches for the prevention of human disease.